The Phene Plate (PhP) system of biochemical fingerprinting of bacteria is a computerised typing system, based on quantitative measurements of the kinetics of several biochemical reactions of bacteria grown in liquid medium in microtitration plates. For each isolate tested, it yields a biochemical fingerprint comprising several kinds of quantitative data which are useful for establishing similarities among strains with a personal-computer program. In this study, a set of 16 specific substrates was chosen to differentiate strains of Salmonella of serotype Typhimurium. The system was evaluated for its typability, reproducibility and discriminatory power in tests with a collection of 100 epidemiologically unrelated Typhimurium strains and results were compared with those obtained by phage typing. At an identity level of 0.980, strains were assigned by this method to 51 biochemical phenotypes (BPTs), giving a diversity index of 0.963 and a resolution index of 0.210. In contrast, 24 phage types (PTs) were identified among these isolates (a diversity index of 0.901). The combined use of biochemical fingerprinting by the PhP system and phage typing discriminated 82 phenotypes (a diversity index of 0.994). Stability of markers in each of the methods was also evaluated after subculture of 20 strains for 21 consecutive days. Only nine biochemical reactions were found that were subject to small, but measurable, changes for at least one isolate. These changes slightly decreased the mean similarity coefficients among strains but the overall BPTs of the strains showed changes in four strains (20 %). In contrast, eight strains (40 %) showed changes in their PTs after this treatment. It is concluded that the PhP system is a highly discriminatory and reproducible method for typing Typhimurium strains. It is easy to perform, and may be used alone or in combination with phage typing in epidemiological studies of Typhimurium strains.